This 5-year project will study interactive signaling between the enteric nervous system (ENS), spinal sensory afferent nerves and enteric mast cells, using the guinea pig small intestine as the experimental model. The project tests a hypothesis, supported by preliminary data, that a positive feed-back signaling loop between spinal afferents, ENS neurons and enteric mast cells amplifies nociceptive and other forms of sensory input from the gut to the central nervous system. The neurophysiology of ENS-spinal afferent-mast cell interactions is basic for understanding visceral hypersensitivity and the emerging recognition that functional abdominal pain can involve co-morbidity of intestinal hypersensitivity with other pain syndromes (e.g., interstitial cystitis, prostatitis, vulvodynia and fibromyalgia). The main objectives are to learn: a) which of the multiple morphological, electrophysiological and immunohistochemical neuronal subtypes in the ENS receive synaptic input from spinal afferents;b) the kinds of synaptic potentials evoked by afferent stimulation in ENS neurons and the identification of the neurotransmitters and receptors involved;c) how afferents innervate enteric mast cells to evoke the release of mast cell mediators and the identity of neurotransmitters released at afferent- mast cell junctions and the post-junctional receptors involved;d) how afferent-evoked release of mast cell mediators influences the electrophysiological and synaptic behavior of identified functional subclasses of neurons in the ENS. The specific aims are: 1) to identify the neurotransmitters released by active intramural spinal afferents at their junctions with single identified ENS neurons and the transmitters released at the junctions of the afferents with intramural mast cells;2) to identify the postsynaptic/postjunctional receptor subtypes that mediate the actions of neurotransmitters released by spinal afferents at synapses on enteric neurons and at junctions with intramural mast cells;3) To identify neurotransmitters released by neurons of the ENS at their junctions with neighboring mast cells and to identify the receptor subtypes for the transmitters, which activate the mast cells to release preformed and lipid membrane-derived paracrine signals.